Frzb is a secreted protein initially isolated from purified cartilage extracts and shares homology to the cysteine rich domain (CRD) of frizzled, a Wnt receptor. Wnt proteins are secreted signaling molecules having numerous developmental functions, including skeletal development, as well as dysfunction in oncogenesis. It was recently shown that Frzb can bind to and inactivate Wnts, leading to speculation for the potential therapeutic use of such activity in modifying Wnt induced developmental and oncogenic events. We have demonstrated that Frzb is temporally and spatially expressed during skeletal and craniofacial development. In an attempt to understand the role of Frzb in skeletal development we are conducting a conditional gene knockout, using the Cre-LoxP recombination system. We are currently analyzing founder mice for germline transmission of the "floxed" Frzb gene. Once confirmed we shall cross the "floxed" mice with a ubiquitously expressing Cre mouse line. In ovo expression of the bacterial enzyme Cre recombinase will result in the excision of the CRD encoding exon resulting in a loss of Frzb function. Should this produce a lethal phenotype, early in development, we will then cross the "floxed" mice with a transgenic line expressing Cre under the control of a limb-specific promoter. Analysis of the regulatory elements of Frzb will provide useful information as to the control of its temporal and spatial expression pattern. We have characterized mouse Frzb and determined its exon/intron structure. Transient transgenic experiments using various promoter sequences are being conducted to determine the location of the tissue-specific regulatory regions.